EP3090757A1 - Mycobactérie recombinée en tant qu'agent d'immunothérapie pour le traitement du cancer - Google Patents

Mycobactérie recombinée en tant qu'agent d'immunothérapie pour le traitement du cancer Download PDF

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EP3090757A1
EP3090757A1 EP15166206.1A EP15166206A EP3090757A1 EP 3090757 A1 EP3090757 A1 EP 3090757A1 EP 15166206 A EP15166206 A EP 15166206A EP 3090757 A1 EP3090757 A1 EP 3090757A1
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cell
recombinant
bcg
rbcg
treatment
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German (de)
English (en)
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Leander Grode
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Vakzine Projekt Management GmbH
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Vakzine Projekt Management GmbH
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Priority to EP15166206.1A priority Critical patent/EP3090757A1/fr
Priority to AU2016258885A priority patent/AU2016258885B2/en
Priority to EP16724315.3A priority patent/EP3291831B1/fr
Priority to KR1020177034007A priority patent/KR102667859B1/ko
Priority to MX2017014142A priority patent/MX2017014142A/es
Priority to CA2984602A priority patent/CA2984602C/fr
Priority to SI201631067T priority patent/SI3291831T1/sl
Priority to CN201680034463.7A priority patent/CN107864656A/zh
Priority to JP2017557195A priority patent/JP6778697B2/ja
Priority to PL16724315T priority patent/PL3291831T3/pl
Priority to ES16724315T priority patent/ES2845724T3/es
Priority to PCT/EP2016/059872 priority patent/WO2016177717A1/fr
Priority to LTEP16724315.3T priority patent/LT3291831T/lt
Priority to EA201792412A priority patent/EA036492B1/ru
Priority to BR112017023747-4A priority patent/BR112017023747A2/pt
Priority to RS20210059A priority patent/RS61333B1/sr
Priority to HUE16724315A priority patent/HUE052711T2/hu
Priority to CUP2017000138A priority patent/CU20170138A7/es
Priority to DK16724315.3T priority patent/DK3291831T3/da
Priority to PT167243153T priority patent/PT3291831T/pt
Priority to US15/571,415 priority patent/US10525118B2/en
Publication of EP3090757A1 publication Critical patent/EP3090757A1/fr
Priority to IL255303A priority patent/IL255303B/en
Priority to PH12017502011A priority patent/PH12017502011A1/en
Priority to HK18107179.8A priority patent/HK1247574A1/zh
Priority to US16/688,371 priority patent/US11426453B2/en
Priority to CY20211100030T priority patent/CY1123754T1/el
Priority to HRP20210107TT priority patent/HRP20210107T1/hr
Priority to US17/667,784 priority patent/US20220160858A1/en
Withdrawn legal-status Critical Current

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/0005—Vertebrate antigens
    • A61K39/0011—Cancer antigens
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/14—Hydrolases (3)
    • C12N9/78—Hydrolases (3) acting on carbon to nitrogen bonds other than peptide bonds (3.5)
    • C12N9/80—Hydrolases (3) acting on carbon to nitrogen bonds other than peptide bonds (3.5) acting on amide bonds in linear amides (3.5.1)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/02—Bacterial antigens
    • A61K39/04—Mycobacterium, e.g. Mycobacterium tuberculosis
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P13/00—Drugs for disorders of the urinary system
    • A61P13/10—Drugs for disorders of the urinary system of the bladder
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y305/00—Hydrolases acting on carbon-nitrogen bonds, other than peptide bonds (3.5)
    • C12Y305/01—Hydrolases acting on carbon-nitrogen bonds, other than peptide bonds (3.5) in linear amides (3.5.1)
    • C12Y305/01005—Urease (3.5.1.5)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
    • A61K2039/52—Bacterial cells; Fungal cells; Protozoal cells
    • A61K2039/522—Bacterial cells; Fungal cells; Protozoal cells avirulent or attenuated
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
    • A61K2039/52—Bacterial cells; Fungal cells; Protozoal cells
    • A61K2039/523—Bacterial cells; Fungal cells; Protozoal cells expressing foreign proteins
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/54—Medicinal preparations containing antigens or antibodies characterised by the route of administration
    • A61K2039/541—Mucosal route
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/545—Medicinal preparations containing antigens or antibodies characterised by the dose, timing or administration schedule
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/58—Medicinal preparations containing antigens or antibodies raising an immune response against a target which is not the antigen used for immunisation
    • A61K2039/585—Medicinal preparations containing antigens or antibodies raising an immune response against a target which is not the antigen used for immunisation wherein the target is cancer

Definitions

  • the invention relates to a recombinant Mycobacterium cell for use as an immunotherapeutic agent in the treatment of cancer, particularly in the treatment of solid tumors. More particularly, the invention relates to the immunotherapy of bladder carcinoma.
  • Urothelial bladder carcinoma is the 5th most common cancer. In the United States, about 75.000 new cases are diagnosed each year 4.5% of all new cancers, and approximately 15.600 deaths are expected. In Germany, about 16.000 new cases are diagnosed each year. Because a recurrence of disease is likely in bladder carcinoma, patients must undergo surveillance for an extended period.
  • bladder carcinomas begin in transitional epithelial cells that make up the inner lining of the bladder. As these tumors grow, they can invade the surrounding connective tissue and muscle. In advanced disease, tumors spread beyond the bladder to nearby lymph nodes or pelvic organs or metastasize to more distant organs such as lung, liver and bone.
  • the overall 5-year survival rate for bladder carcinoma is 77%, and this rate has not changed significantly over the last 10 years.
  • the 5-year relative survival rates for patients with tumors restricted to the inner layer of the bladder are 96% and 69%, respectively.
  • the rates drop to 34% for those with disease that has spread locally beyond the bladder and to 6% with distant metastases.
  • BCG Bacillus Calmette Guérin
  • Standard treatment for patients with muscle-invasive bladder carcinoma includes cisplatin-based chemotherapy followed by surgical removal of the bladder or radiation therapy and concomitant chemotherapy.
  • Recurrent bladder carcinoma may be treated with combination therapy regimens, including gemcitabine plus cisplatin or methotrexate, vinblastine, doxorubicin plus cisplatin.
  • a recombinant BCG strain expressing a phagolysosomal escape domain is described in WO 99/101496 , the content of which is herein incorporated by reference.
  • the phagolysosomal escape domain enables the strain to escape from the phagosome of infected host cells by perforating the membrane of the phagosome.
  • a urease-deficient recombinant strain was developed. This strain is disclosed in WO2004/094469 , the content of which is herein incorporated.
  • WO 2012/085101 discloses that a recombinant ⁇ ureC Hly + rBCG (rBCG) strain expressing membrane-perforating listeriolysin (Hly) of Listeria monocytogenes and devoid of urease C induces superior protection against aerogenic challenge with Mycobacterium tuberculosis (MTB) as compared to parental BCG (pBCG) in a preclinical model. Further, it is shown that rBCG and pBCG induce marked Th1 immune responses, whilst only rBCG elicits are profound Th17 response in addition.
  • MTB Mycobacterium tuberculosis
  • a subject-matter of the present invention is a recombinant Mycobacterium cell which comprises a recombinant nucleic acid molecule encoding a fusion polypeptide comprising:
  • a further aspect of the present invention is a method for the immunotherapeutic treatment of solid tumors in a subject in need thereof, comprising administering to said subject a recombinant Mycobacterium cell which comprises a recombinant nucleic acid molecule encoding a fusion polypeptide comprising:
  • vesicular instillation of a recombinant BCG cell into the bladder of rats surprisingly results in an increased infiltration of urinary bladder tissue by lymphocytes, particularly CD4- and CD8-positive lymphocytes resulting in a high incidence of focal and/or multifocal lymphocytic infiltration.
  • lymphocytes particularly CD4- and CD8-positive lymphocytes
  • the urinary bladder tissue of animals treated with standard BCG showed CD4- and CD8-positive lymphocytes only as single cell infiltrate levels (diffuse infiltration).
  • administration of a recombinant BCG cell did not raise any safety issues.
  • the present invention also relates to a recombinant Mycobacterium cell as described above for use as an immunotherapeutic agent in the treatment of solid tumors in order to obtain focal and/or multifocal lymphocytic infiltration, e.g. with CD4 and CD8 T cells at the site of administration.
  • the recombinant Mycobacterium cell of the present invention is particularly suitable for use in human medicine.
  • the immunotherapeutic agent is a live recombinant Mycobacterium cell which comprises a recombinant nucleic acid molecule encoding a fusion polypeptide comprising (a) a domain capable of eliciting an immune response and (b) a phagolysosomal escape domain.
  • the domain capable of eliciting an immune response is preferably an immunogenic peptide or polypeptide from a pathogen or an immunogenic fragment thereof.
  • the Mycobacterium cell is preferably an M. bovis cell, an M. tuberculosis cell, particularly an attenuated M. tuberculosis cell or other Mycobacteria, e.g. M. microti , M. smegmatis , M. canettii , M. marinum or M. fortuitum . More preferably, the cell is an attenuated recombinant M . bovis (BCG) cell, particularly a M. bovis BCG cell, more particularly a recombinant M . bovis BCG cell from strain Danish subtype Prague ( Brosch et al., Proc. Natl. Acad. Sci. USA, 104 (2007), 5396-5601 ).
  • BCG attenuated recombinant M . bovis
  • the Mycobacterium cell is recombinant urease-deficient.
  • the ureC sequence of the Mycobacterium cell is inactivated ( ⁇ Urec), e.g. by constructing a suicide vector containing a ureC gene disrupted by a selection marker gene, e.g. the hygromycin gene, transforming the target cell with the vector and screening for selection marker-positive cells having a urease negative phenotype.
  • the selection marker gene i.e. the hygromycin gene, is subsequently inactivated.
  • the cell is a selection marker-free recombinant Mycobacterium cell.
  • the cell is selection marker-free recombinant BCG strain Danish subtypevic characterized as rBCG ⁇ Urec :: Hly + .
  • the domain capable of eliciting an immune response is preferably selected from immunogenic peptides or polypeptides from M. bovis, M. tuberculosis or M. leprae or from immunogenic fragments thereof having a length of at least 6, preferably at least 8 amino acids, more preferably at least 9 amino acids and e.g. up to 20 amino acids.
  • suitable antigens are Ag85B (p30) from M. tuberculosis, Ag85B ( ⁇ -antigen) from M. bovis BCG, Ag85A from M. tuberculosis and ESAT-6 from M. tuberculosis and fragments thereof.
  • the domain capable of eliciting an immune response is selected from non-Mycobacterium polypeptides.
  • the immunogenic domain is derived from the antigen Ag85B. Most preferably, the immunogenic domain comprises the sequence from aa.41 to aa.51 in SEQ ID No.2.
  • the recombinant nucleic acid molecule further comprises a phagolysosomal escape domain, i.e. a polypeptide domain which provides for an escape of the fusion polypeptide from the phagolysosome into the cytosol of mammalian cells.
  • a phagolysosomal escape domain i.e. a polypeptide domain which provides for an escape of the fusion polypeptide from the phagolysosome into the cytosol of mammalian cells.
  • the phagolysosomal escape domain is a Listeria phagolysosomal escape domain, which is described in US 5,733,151 , herein incorporated by reference. More preferably, the phagolysosomal escape domain is derived from the listeriolysin gene (Hly) of L.monocytogenes.
  • the phagolysosomal domain is encoded by a nucleic acid molecule selected from: (a) a nucleotide sequence comprising nucleotides 211 - 1722 as shown in SEQ ID No.1, (b) a nucleotide sequence which encodes for the same amino acid sequence as the sequence from (a), and (c) a nucleotide sequence hybridizing under stringent conditions with the sequence from (a) or (b).
  • a nucleic acid molecule selected from: (a) a nucleotide sequence comprising nucleotides 211 - 1722 as shown in SEQ ID No.1, (b) a nucleotide sequence which encodes for the same amino acid sequence as the sequence from (a), and (c) a nucleotide sequence hybridizing under stringent conditions with the sequence from (a) or (b).
  • the present invention also comprises nucleic acid sequences hybridizing therewith.
  • hybridization is used as defined in Sambrook et al. (Molecular Cloning. A laboratory manual, Cold Spring Harbor Laboratory Press (1989), 1.101-1.104 ).
  • hybridization is used if a positive hybridization signal can still be observed after washing for one hour with 1 X SSC and 0.1 % SDS at 55°C, preferably at 62° C and more preferably at 68°C, particularly for 1 hour in 0.2 X SSC and 0.1 % SDS at 55°C, preferably at 62°C and more preferably at 68°C.
  • a sequence hybridizing with a nucleotide sequence as per SEQ ID No.1 under such washing conditions is a phagolysosomal escape domain encoding nucleotide sequence preferred by the subject invention.
  • a nucleotide sequence encoding a phagolysosomal escape domain as described above may be directly obtained from a Listeria organism or from any recombinant source e.g. a recombinant E.coli cell containing the corresponding Listeria nucleic acid molecule or a variant thereof as described above.
  • the recombinant nucleic acid molecule encoding for a fusion polypeptide contains a signal peptide encoding sequence.
  • the signal sequence is a signal sequence active in Mycobacteria, preferably in M.bovis, e.g. a native M.bovis signal sequence.
  • a preferred example of a suitable signal sequence is the nucleotide sequence coding for the Ag85B signal peptide which is depicted in SEQ ID No.1 from nucleotide 1 to 120.
  • a peptide linker be provided between the immunogenic domain and the phagolysosomal escape domain.
  • said peptide linker has a length of from 5 to 50 amino acids. More preferably, a sequence encoding a linker as shown in SEQ ID No.1 from nucleotide 154 to 210 or a sequence corresponding thereto as regards the degeneration of the genetic code.
  • the nucleic acid may be located on a recombinant vector.
  • the recombinant vector is a prokaryotic vector, i.e. a vector containing elements for replication or/and genomic integration in prokaryotic cells.
  • the recombinant vector carries the nucleic acid molecule of the present invention operatively linked with an expression control sequence.
  • the expression control sequence is preferably an expression control sequence active in Mycobacteria, particularly in M.bovis.
  • the vector can be an extrachromosomal vector or a vector suitable for integration into the chromosome. Examples of such vectors are known to the man skilled in the art and, for instance, given in Sambrook et al . supra.
  • the immunotherapeutic agent of the present invention is suitable for the treatment of solid tumors, such as bladder, lung, liver, breast, kidney or prostate tumors.
  • solid tumors such as bladder, lung, liver, breast, kidney or prostate tumors.
  • the present invention is suitable for the treatment of non-invasive solid tumors.
  • the solid tumor is bladder carcinoma, e.g., non-invasive bladder carcinoma, e.g.
  • non-invasive papillary carcinoma in situ Ta
  • non-invasive carcinoma in situ Tcis
  • tumor invading subepithelial connective tissue T 1
  • tumor invading superficial muscle inner half
  • tumor invading deep muscle outer half
  • tumor invading perivesical tissue T3 including T3a and T3b
  • tumor invading prostate uterus or vagina
  • tumor invading pelvic wall or abdominal wall T4b
  • the tumor is a superficial tumor or carcinoma in situ (Tcis), non-invasive papillary carcinoma (Ta), or a tumor invading subepithelial connective tissue (T1).
  • the immunotherapeutic treatment is suitable for the treatment of primary tumors and/or for the treatment of recurring tumors.
  • the immunotherapeutic agent is preferably locally administered to the tumor site, i.e., to the site of a primary tumor before surgery or after surgery and optionally after chemotherapy.
  • the agent is preferably administered by vesicular instillation into the urinary bladder.
  • the administration may involve local injection or, in case of lung tumors, pulmonal administration.
  • the immunotherapeutic agent of the invention may be administered as a first-line immunotherapy in patients, who have not been treated previously with an anti-tumor-immunotherapeutic agent such as standard BCG, or a follow-up immunotherapy in patients who have been previously treated with anti-tumor-immunotherapeutic agent such as standard BCG.
  • the immunotherapeutic agent may be administered with a newly diagnosed solid tumor, e.g., a bladder carcinoma, or to patients, particularly patients with recurrent solid tumors, e.g., bladder carcinoma.
  • the immunotherapeutic agent is administered to the subject to be treated in an effective dose.
  • the dose for an administration may be about 10 7 to 10 10 viable units, e.g., 10 8 to 10 9 viable units.
  • the immunotherapeutic aget is administered several times, e.g. at least 5 times up to 30 times at predetermined times during the treatment.
  • the immunotherapeutic agent is usually provided as a pharmaceutical preparation which comprises the recombinant Mycobacterial cell in solid form, e.g., a lyophilized or cryoconserved preparation, which is reconstituted with a suitable liquid carrier before use.
  • a suitable liquid carrier e.g., water, water, or water.
  • the preparation may be provided in liquid form, e.g., as suspension.
  • the immunotherapeutic agent of the invention is administered for the treatment of carcinoma in situ.
  • a standard schedule may comprise weekly administration of the agent for at least 4, e.g., 4, 5, 6, 7 or 8 weeks as an induction therapy.
  • the induction therapy should not start until 2-3 weeks after primary tumor surgery.
  • administration may continue using maintenance therapy for at least 6 months or at least 1 year.
  • the immunotherapeutic agent is administered in an induction therapy in the prophylactic treatment of tumor recurrence.
  • therapy may start about 2-3 weeks after biopsy of the tumor site and be repeated, e.g., at weekly intervals for at least 4, e.g., 4, 5, 6, 7 or 8 weeks. In intermediate and high-risk tumors this may be followed by maintenance therapy.
  • Maintenance therapy may comprise long-term therapy, e.g., 6, 9 or 12 months therapy or even longer with treatments at monthly intervals.
  • maintenance therapy may comprise 2, 3 or 4 administrations at weekly intervals, at month 3, 6, 12, 18, 24, 30 and 36.
  • the administration as immunotherapeutic agent of the recombinant Mycobacterium cell to site of a solid tumor as described above may be combined with further anti-tumor therapy, e.g., radiation and/or chemotherapy.
  • the immunotherapy as described above may be combined with a non-tumor site specific administration of the recombinant Mycobacterium cell in order to provide a general stimulation of the immune system.
  • This non-site specific administration may be effected as described in WO 2012/085101 , e.g. before surgery of the primary tumor.
  • the agent is preferably administered to a human subject in a dose of about 1-10 x 10 5 , preferably about 2-8 x 10 5 cells.
  • the agent is preferably administered as a single dose, e.g., by injection. Subcutaneous injection is preferred. Further it is preferred to administer the agent without adjuvant.
  • Dose regime Group 1 Control (diluent) Group 2: ⁇ 2 ⁇ 10 6 rBCG Group 3: (lyophilized)/animal ⁇ 2 ⁇ 10 8 rBCG (lyophilized)/animal Group 4: ⁇ 2 ⁇ 10 6 rBCG (frozen w/o cryoprotectant)/ animal Group 5: ⁇ 2 ⁇ 10 6 CFU BCG medac/animal Route of administration Intravesical instillation in the bladder Frequency of administration Single dose on test day 1. Administration volume 500 ⁇ L/animal Duration of study • 12 adaptation days • 4 in-life test weeks • 28 incubation days 1.2 Results Mortality None of the animals died prematurely.
  • BCG medac group 5
  • control group 1
  • No reference-item related CFU counts were noted for the examined organs and the blood of the animals treated once with an intravesical instillation of 2x10 6 BCG medac/animal.
  • no CFU counts at all were noted for the urinary bladder four weeks after instillation of the reference item, indicating a rapid clearance of the administered mycobacteria from the site of instillation.
  • No differences were noted between the animals treated with 2x10 6 BCG medac / animal and the control animals.
  • rBCG groups 2 to 4
  • Immunohistochemistry for lymphocyte sub-typing in the urinary bladder tissue revealed the highest incidence of focal and multifocal lymphocytic infiltration with CD4 and CD8 positive cells in groups 2 and 3 treated once with 2x10 6 or 2x10 8 CFU rBCG (lyophilized)/animal by intravesical instillation, whereas animals of groups 1 (control), 4 and 5 expressed CD4 and CD8 positive lymphocytes only as single cell infiltrate levels (diffuse infiltration). Nearly all animals (12 of 13) of groups 1 (control), 4 and 5 expressed CD4 and CD8 positive lymphocytes only as single cell infiltrate levels (diffuse infiltration).
  • Immunohistochemistry for lymphocyte sub-typing in the urinary bladder tissue revealed the highest incidence of focal and multifocal lymphocytic infiltration with CD4 and CD8 positive cells in groups 2 and 3 treated once with 2x10 6 or 2x10 8 CFU rBCG/animal by intravesical instillation, whereas animals of groups 1 (control), 4 and 5 expressed CD4 and CD8 positive lymphocytes only as single cell infiltrate levels (diffuse infiltration). Nearly all animals (12 of 13) of groups 1 (control), 4 and 5 expressed CD4 and CD8 positive lymphocytes only as single cell infiltrate levels (diffuse infiltration).
  • Fig. 1A shows focal and multifocal lymphocytic infiltration after administration of rBCG.
  • Fig. 1B shows only diffuse and singular infiltration after administration of BCG medac (200x magnification).
  • Dose regime Group 1 Control (diluent) Group 2: ⁇ 2 ⁇ 10 6 CFU rBCG (lyophilized)/animal Group 3: ⁇ 2 ⁇ 10 8 CFU rBCG (lyophilized)/animal Group 4: ⁇ 2 ⁇ 10 6 CFU rBCG (frozen w/o cryoprotectant)/animal Group 5: ⁇ 2 ⁇ 10 6 CFU BCG medac/animal Route of administration Intravesical instillation in the bladder Frequency of administration Repeated administration; once weekly on test days 1, 8, 15, 22, 29, and 36. Administration volume 500 ⁇ L/animal Duration of study • 21 adaptation days • 9 in-life test weeks • 28 incubation days 2.2 Results Mortality None of the animals died prematurely.
  • CFU counts rBCG (groups 2 to 4) vs. control (group 1)
  • No test-item related CFU counts were noted for the examined organs and the blood of the animals treated six times with an intravesical instillation of 2 ⁇ 10 6 or 2 ⁇ 10 8 CFU rBCG (lyophilized)/animal, or of 2 ⁇ 10 6 CFU rBCG (frozen/w/o cryoprotectant)/animal.
  • no CFU counts at all were noted for the urinary bladder four weeks after the last instillation of the test item, indicating a rapid clearance of the administered mycobacteria from the site of instillation.
  • Immunohistochemistry for lymphocyte sub-typing in the urinary bladder tissue revealed the highest incidence of multifocal lymphocytic infiltration with CD4 and CD8 positive cells in group 2 treated with 2 ⁇ 10 6 CFU rBCG/animal by 6 intravesical instillations, followed by group 3 treated with 2 ⁇ 10 8 CFU rBCG/animal by 6 intravesical instillations, whereas animals of groups 1 (control), 4 and 5 expressed CD4 and CD8 positive lymphocytes only as single cell infiltrate levels (diffuse infiltration). Neutrophilic granulocytes could rarely be detected in any slides examined.
  • Fig. 2A shows focal and multifocal lymphocytic infiltration after administration of rBCG.
  • Fig. 2B shows only diffuse and singular infiltration after administration of BCG medac (200x magnification).

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EP15166206.1A 2015-05-04 2015-05-04 Mycobactérie recombinée en tant qu'agent d'immunothérapie pour le traitement du cancer Withdrawn EP3090757A1 (fr)

Priority Applications (28)

Application Number Priority Date Filing Date Title
EP15166206.1A EP3090757A1 (fr) 2015-05-04 2015-05-04 Mycobactérie recombinée en tant qu'agent d'immunothérapie pour le traitement du cancer
EA201792412A EA036492B1 (ru) 2015-05-04 2016-05-03 Рекомбинантная клетка mycobacterium bovis как иммунотерапевтическое средство для лечения рака мочевого пузыря
HUE16724315A HUE052711T2 (hu) 2015-05-04 2016-05-03 Rekombináns Mycobaktérium immunterápiás szerként rák kezelésére
RS20210059A RS61333B1 (sr) 2015-05-04 2016-05-03 Rekombinantna mycobacterium kao imunoterapeutski agens za tretiranje kancera
MX2017014142A MX2017014142A (es) 2015-05-04 2016-05-03 Mycobacterium recombinante como un agente inmunoterapeutico para el tratamiento de cancer.
EP16724315.3A EP3291831B1 (fr) 2015-05-04 2016-05-03 Mycobactérie de recombinaison en tant qu'agent immunothérapeutique pour le traitement du cancer
SI201631067T SI3291831T1 (sl) 2015-05-04 2016-05-03 Rekombinantni mycobacterium kot imunoterapevtsko sredstvo za zdravljenje raka
CN201680034463.7A CN107864656A (zh) 2015-05-04 2016-05-03 重组分枝杆菌作为免疫治疗剂用于治疗癌症
JP2017557195A JP6778697B2 (ja) 2015-05-04 2016-05-03 癌の治療のための免疫療法薬としての、組み換えマイコバクテリウム
PL16724315T PL3291831T3 (pl) 2015-05-04 2016-05-03 Rekombinowana mykobakteria jako środek immunoterapeutyczny do leczenia nowotworu
ES16724315T ES2845724T3 (es) 2015-05-04 2016-05-03 Micobacteria recombinante como agente inmunoterapéutico para el tratamiento de cáncer
PCT/EP2016/059872 WO2016177717A1 (fr) 2015-05-04 2016-05-03 Mycobactérie de recombinaison en tant qu'agent immunothérapeutique pour le traitement du cancer
CUP2017000138A CU20170138A7 (es) 2015-05-04 2016-05-03 Mycobacterium recombinante como un agente inmunoterapéutico para el tratamiento de cáncer
AU2016258885A AU2016258885B2 (en) 2015-05-04 2016-05-03 Recombinant Mycobacterium as an immunotherapeutic agent for the treatment of cancer
BR112017023747-4A BR112017023747A2 (pt) 2015-05-04 2016-05-03 micobactéria recombinante como um agente imunoterapêutico para o tratamento de câncer
KR1020177034007A KR102667859B1 (ko) 2015-05-04 2016-05-03 암 치료용 면역요법제로서의 재조합 미코박테리움
CA2984602A CA2984602C (fr) 2015-05-04 2016-05-03 Mycobacterie de recombinaison en tant qu'agent immunotherapeutique pour le traitement du cancer
LTEP16724315.3T LT3291831T (lt) 2015-05-04 2016-05-03 Rekombinantinė bakterija kaip imunoterapinis agentas, skirtas vėžio gydymui
DK16724315.3T DK3291831T3 (da) 2015-05-04 2016-05-03 Rekombinant mykobakterie som et immunterapeutisk middel til behandling af cancer
PT167243153T PT3291831T (pt) 2015-05-04 2016-05-03 Mycobacterium recombinante como um agente imunoterapêutico para o tratamento de cancro
US15/571,415 US10525118B2 (en) 2015-05-04 2016-05-03 Recombinant Mycobacterium as an immunotherapeutic agent for the treatment of cancer
IL255303A IL255303B (en) 2015-05-04 2017-10-29 Recombinant mycobacterium as an immunotherapeutic agent for cancer treatment
PH12017502011A PH12017502011A1 (en) 2015-05-04 2017-11-03 Recombinant mycobacterium as an immunotherapeutic agent for the treatment of cancer
HK18107179.8A HK1247574A1 (zh) 2015-05-04 2018-05-31 重組分枝桿菌作為免疫治療劑用於治療癌症
US16/688,371 US11426453B2 (en) 2015-05-04 2019-11-19 Recombinant mycobacterium as an immunotherapeutic agent for the treatment of cancer
CY20211100030T CY1123754T1 (el) 2015-05-04 2021-01-15 Ανασυνδιασμενο mycobacterium ως ενας ανοσοθεραπευτικος παραγοντας για τη θεραπεια του καρκινου
HRP20210107TT HRP20210107T1 (hr) 2015-05-04 2021-01-21 Rekombinantna mikobakterija kao imunoterapijski agens za liječenje raka
US17/667,784 US20220160858A1 (en) 2015-05-04 2022-02-09 Recombinant Mycobacterium as an Immunotherapeutic Agent for the Treatment of Cancer

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US10525118B2 (en) 2020-01-07
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CA2984602A1 (fr) 2016-11-10
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SI3291831T1 (sl) 2021-03-31
US11426453B2 (en) 2022-08-30
US20180200357A1 (en) 2018-07-19
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AU2016258885B2 (en) 2021-12-09
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HK1247574A1 (zh) 2018-09-28
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KR102667859B1 (ko) 2024-05-21
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